Menu
Institut de Biologie StructuraleGrenoble / France

Contact person(s) related to this article / BOISBOUVIER Jerome / FAVIER Adrien

NMR of Large Biomolecular Assemblies

Go Back To

Team Members

Jérôme Boisbouvier (CNRS Researcher)
Pierre Gans (CEA Researcher)
Lionel Imbert (CNRS Engineer)
Faustine Henot (PhD)
Ricarda Törner (PhD)

If you want to contact team members, please use the generic e-mail address: 
firstname(dot)lastname(at)ibs(dot)fr

Presentation

Our team is synergistically developing advanced in vivo and in vitro labeling methods together with optimized NMR spectroscopy to push forward the limit of application of NMR into Biology. We are integrating these innovative methods with other biophysics approaches in order to study folding, structure, dynamics, transient interactions and mechanism of essential cellular machineries and large chaperones

Recent Applications:

Self-assembling of the 468kDa TET2 proteolytic machine.


We have combined methyl specific isotopic labeling with relaxation-optimized NMR spectroscopy to investigate in real-time the self-assembly process of the 468 kDa TET2 proteolytic machinery at the structural level, including the kinetics, thermodynamics and molecular shapes of intermediate states, integrating also time-resolved EM and mass spectrometry (Macek et al. Science Advances 2017)

Structure determination of a 0.5 MDa complex using NMR and cryo-EM.


In this study, we combined the possibility to extract by NMR, secondary structure information and long range distance restraints, enabled by specific isotope-labeling schemes, together with cryo-EM map in order to solve the structure of the 12 x 39 kDa-large TET2 assembly to a precision below 1 Å (Gauto et al. Nature communication 2019). This new approach provides high precision structures even in case where only medium to low resolution cryo-EM density maps are available.

Structural and Functional Study of a 1 MDa Chaperonin in Action.


In this project, we have demonstrated how solution NMR can provide functional insight into the 1 MDa large hsp60 chaperonin, while the machinery is processing client proteins. We reveal how nucleotide binding, hydrolysis and release control switching between closed and open states (Mas et al. Science Advances 2018). This approach opens new perspectives to study structures and mechanisms of various ATP-driven biological machineries in the heat of action.

The team is operating the ISBG Cell-Free Facility offering the possibility to visiting scientists to express in large scale target proteins for structural biology investigation. For more information visit cell-free facility page here

Major Publications:

• Van Melckebeke H, Devany M, Di Primo C, Beaurain F, Toulmé JJ, Bryce DL, Boisbouvier J. “Liquid Crystal NMR Structure of HIV TAR RNA Bound to its SELEX RNA Aptamer Reveals the Origins of the High Stability of the Complex“ Proc. Natl. Acad. Sci. USA 105, 9210-5 (2008).

• Amero C, Schanda P, Dura A, Ayala I, Marion D, Franzetti B, Brutscher B, Boisbouvier J. “Fast Two Dimensional NMR Spectroscopy of High Molecular Weight Protein Assemblies”. J. Am. Chem. Soc. 131, 3448-9 (2009).

• Gans P, Hamelin O, Sounier R, Ayala I, Dura MA, Amero C, Noirclerc-Savoye M, Franzetti B, Plevin MJ, Boisbouvier J. “Stereospecific Isotopic Labeling of Methyl Groups for NMR Studies of High Molecular Weight Proteins” Angew. Chem. Int. Ed. 49, 1958-1862 (2010).

• PlevinMJ, Bryce DL, Boisbouvier J. “Direct Detection of CH••Pi Interactions in Proteins” Nature Chemistry. 2, 466-471 (2010).

• Kerfah R, Plevin MJ, Sounier R., Gans P, Boisbouvier J. “Methyl Specific Isotopic Labeling: A Molecular Tool Box for NMR Studies of Large Proteins“ Current Opinion in Structural Biology 32, 113-122 (2015).

• Macek P, Kerfah R, Boeri Erba E, Crublet E, Moriscot C, Schoehn G, Amero C, Boisbouvier J. “Unraveling Self-Assembly Pathways of the 468 kDa Proteolytic Machine TET2” Science Advances 3, e1601601 (2017).

• Mas G, Guan J-Y, Crublet E, Colas Debled E, Moriscot C, Gans P, Schoehn G, Macek P, Schanda P, Boisbouvier J. “Structural Investigation of a Chaperonin in Action Reveals How Nucleotide Binding Regulates the Functional Cycle“ Science Advances 4, eaau4196 (2018).

• Gauto D, Estrozi L, Schwieters C, Effantin G, Macek P, Sounier R, Sivertsen AC, Schmidt E, Kerfah R, Mas G, Colletier JP, Güntert P, Favier A, Schoehn G, Schanda P, Boisbouvier J. “Integrated NMR and cryo-EM atomic-resolution structure determination of a half-megadalton enzyme complex“ Nature Communication 10(1):2697 (2019).

Academic collaborations:

ForschungsZentrum-Jülich (L. Gremer; W. Hoyer; D. Willbold); HITS-Heildelberg (D. Kokh; K. Sadiq; R. Wade); IBS (B. Brutscher; P. Schanda; G. Schoehn); ILM-Lyon (C. Loison); Sanofi (O. Frances); University P. & M. Curie - Paris (E. Miclet).

Funding Agencies: